. MRF4 is a regulatory factor important for vertebrate skeletal muscle determination and differentiation and may mediate the effects of innervation on myofiber gene expression. Understanding how muscle cells are determined and differentiate will therefore require understanding how MRF4 expression is regulated. Towards this end, the principal investigator proposes: (1) to identify the cis-regulatory elements of the Xenopus MRF4 (XMRF4) gene. Preliminary work has shown that the regulatory region of XMRF4 differs from that of known mammalian MRF4s. Deletion mutants of the XMRF4 promoter region will be fused to LAC-Z or GFP reporter plasmids and injected into zygotes. Expression of the foreign genes will be monitored in embryonic muscle by in situ hybridization, fluorescence, or chemiluminescence.(2) Identify trans-activators of XMRF4 , especially to determine whether the other four known muscle regulatory factors trans-activate XMRF4 when they are overexpressed in non-muscle cells from Xenopus embryos or from tissue culture. (3) Determine whether expression of XMRF4 differs in experimentally aneural muscle cells, and (4) Create transgenic Xenopus with stably integrated XMRF4 constructs for future studies of XMRF4 regulation.